The goal of this research is to contribute to an understanding of the structural and functional organization of the eukaryotic chromosome, especially in relation to the process of DNA replication. Yeast, a simple eukaryote, will be used as a model system. The nature of initiation sites for DNA replication will be studied in both the cytoplasmic 2-micron DNA and the chromosomal genes coding for ribosomal RNA (rRNA). These experiments will determine if initiation occurs at specific loci in these two DNAs. The molecular structure of initiation sites and controls over their activation will also be investigated. In addition, studies to determine the number and size of blocks of rRNA genes will be pursued. Finally, proteins with possible roles in DNA replication will be isolated by their affinity for DNA. The biological role of these proteins will be determined by investigating their characteristics in extracts from mutants temperature sensitive for DNA synthesis.